In this research proposal we plan to apply biochemical and genetic approaches to the study of amino acid transport in eukaryotic cells. We are characterizing the transport systems for neutral amino acids. 3T3 mouse cells, baby hamster kidney cells, Chinese hamster ovary cells and Ehrlich ascites tumor cells will be used. Membrane vesicle preparations will be made from the cells and used for transport studies. We will continue our studies on the use of Triton X-100 to solubilize the membranes and isolate leucine-binding activity from Ehrlich ascites tumor cells. This binding activity for leucine resembles that of the L transport system which we previously described. Reconstitution of leucine transport in phospholipid vesicles will also be attempted. Another aim of the proposal is the study of amino acid regulation in normal and virus-transformed cells. We hope to determine if membrane transport changes occur during malignant transformation and if so whether they are early or late events in the transformation process. In these studies we will measure transport changes in the primary cell of rat epidermal cells as they are being transformed by chemical carcinogens in vitro. Finally we plan to select transport mutants in Chinese hamster ovary cells and genetically map potential mutants to human chromosomes by cell fusion techniques forming human-hamster cell hybrids.